Programmed death-1 (PD-1) appears transiently on the surface of T cells following immune activation and is highly expressed in T cells chronically exposed to antigen. While PD-1 likely plays an important role in governing responses to self antigen, there is compelling evidence that PD-1-mediated inhibitory signals also play a central role in the functional exhaustion of T cells during chronic viral infection. This has been demonstrated in several animal and human viral infections including HIV. Most notably, antibody blockade between PD-1 and its ligands results in reactivation of the exhausted antigen-specific T cells. This effect has been demonstrated in a number of persistent viral infections, including those from HIV in humans and in the mouse with lymphocytic choriomenigitis virus (LCMV). These data strongly suggest that PD-1 is more than just a marker of T cell exhaustion but rather a critical mediator of this phenotype. These data also imply that manipulating PD-1 expression, signaling, and/or function could lead to novel immune based therapies to treat chronic infection. Surprisingly, virtually nothing is known about the regulation of PD-1 gene expression at the molecular level this basic tenet of PD-1 biology and immunology will be elucidated in this proposal. Our preliminary data provide evidence that PD-1 Is regulated at multiple levels. Using comparative genomics, DNasel hypersensitivity assays, gene reporter assays, and bisulfite sequence, we have identified regions in both mouse and human genomes that are likely to be involved in the regulation of PD-1 and have correlated two of these regions with transcriptional activity. We have also found evidence that the PD-1 gene is differentially methylated in viral-specific CDS T cells in vivo, suggesting a role for epigenetic control of the gene. Experiments in this application will identify the cis-acting elements, Aim 1; determine the breadth and dynamics of epigenetic mechanisms, Aim 2; and identify transcription factors that are associated with PD-1 gene regulation, Aim 3. In all of the aims we will begin with model cell lines, confirm and verify that information in primary mouse and human CDS T cells, test the validity .of these findings in antigen-specific CDS T cell following LCMV infection; and examine HIV-specific CDS T cells to determine how PD-1 regulation factors into HIV disease progression. These approaches will provide molecular targets for pathways that may be ultimately used for treating HIV disease, chronic infection, autoimmunity and aiding to transplantation success.